IgE antibodies bind the high affinity IgE Fc receptor (Fc?RI), found primarily on mast cells and basophils, and trigger inflammatory cascades of the allergic response. Inhibitors of IgE:Fc?RI binding have been identified and an anti-IgE therapeutic antibody (omalizumab) is used to treat severe allergic asthma. However, preformed IgE:Fc?RI complexes that prime cells prior to allergen exposure dissociate extremely slowly and cannot be disrupted by classical competitive inhibitors. We demonstrated that two DARPIn inhibitors not only block IgE:Fc?RI interactions, but they actively disrupt preformed IgE:receptor complexes by a mechanism known as facilitated dissociation. The results indicate that IgE-Fc conformational fluctuations and states, alone and in complexes with Fc?RI, can be targeted by novel, non-classical inhibitors that can actively disassemble IgE:Fc?RI complexes to block the allergic response. In this proposal, we will explore the mechanism of inhibition and facilitated dissociation of IgE:Fc?RI complexes by the DARPins and omalizumab in order to better define the major parameters that govern the efficiency of disruptive inhibition and receptor complex dissociation. The proposed experiments will provide greater insight into the dynamics of IgE:Fc?RI complexes, stabilization of receptor complexes mediated by the IgE-Fc C?2 domains and provide a foundation for designing improved disruptive inhibitors with greater potency and efficiency of complex disassembly.